Stability and Reproducibility Underscore Utility of RT-QuIC for Diagnosis of Creutzfeldt-Jakob Disease

Mol Neurobiol. 2016 Apr;53(3):1896-1904. doi: 10.1007/s12035-015-9133-2. Epub 2015 Apr 1.

Abstract

Real-time quaking-induced conversion (RT-QuIC) allows the amplification of miniscule amounts of scrapie prion protein (PrP(Sc)). Recent studies applied the RT-QuIC methodology to cerebrospinal fluid (CSF) for diagnosing human prion diseases. However, to date, there has not been a formal multi-centre assessment of the reproducibility, validity and stability of RT-QuIC in this context, an indispensable step for establishment as a diagnostic test in clinical practice. In the present study, we analysed CSF from 110 prion disease patients and 400 control patients using the RT-QuIC method under various conditions. In addition, "blinded" ring trials between different participating sites were performed to estimate reproducibility. Using the previously established cut-off of 10,000 relative fluorescence units (rfu), we obtained a sensitivity of 85% and a specificity of 99%. The multi-centre inter-laboratory reproducibility of RT-QuIC revealed a Fleiss' kappa value of 0.83 (95% CI: 0.40-1.00) indicating an almost perfect agreement. Moreover, we investigated the impact of short-term CSF storage at different temperatures, long-term storage, repeated freezing and thawing cycles and the contamination of CSF with blood on the RT-QuIC seeding response. Our data indicated that the PrP(Sc) seed in CSF is stable to any type of storage condition but sensitive to contaminations with blood (>1250 erythrocytes/μL), which results in a false negative RT-QuIC response. Fresh blood-contaminated samples (3 days) can be rescued by removal of erythrocytes. The present study underlines the reproducibility and high stability of RT-QuIC across various CSF storage conditions with a remarkable sensitivity and specificity, suggesting RT-QuIC as an innovative and robust diagnostic method.

Keywords: Cerebrospinal fluid; Creutzfeldt-Jakob disease; Diagnostic test; Prion protein; Real-time quaking-induced conversion.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adolescent
  • Adult
  • Aged
  • Aged, 80 and over
  • Biological Assay / methods*
  • Creutzfeldt-Jakob Syndrome / blood
  • Creutzfeldt-Jakob Syndrome / cerebrospinal fluid
  • Creutzfeldt-Jakob Syndrome / diagnosis*
  • Female
  • Humans
  • Male
  • Middle Aged
  • Preservation, Biological
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Time Factors
  • Young Adult
  • tau Proteins / metabolism

Substances

  • tau Proteins